Engrailed 2 serves as a master regulator of the super-enhancer in the TNC gene locus in non-small cell lung cancer.

Engrailed 2 (EN2) is a homeodomain-containing protein that is dysregulated in many types of cancer. However, the role of EN2 in non-small cell lung cancer (NSCLC) and the mechanism underlying its biological function are largely unclear. Here, we showed that EN2 played an oncogenic function in NSCLC and greatly enhanced the malignant phenotype of NSCLC cells. Meanwhile, EN2 was able to boost the expression of a well-studied oncogenic Tenascin-C (TNC) gene, which in turn activated the AKT signaling pathway. Interestingly, we found that EN2 directly bound to the super-enhancer (SE) region in the TNC locus. The histone marker H3K27ac was also enriched in the region, indicating the activation of the SE. Treatment of the cells with JQ1, an inhibitor of SE activity, abrogated the effect of EN2 on the expression of TNC and phosphorylation of AKT-Ser473. Collectively, our work unveils a novel mode of EN2 function, in which EN2 governs the SE in the TNC locus, consequently activating the oncogenic TNC-AKT axis in NSCLC.

A novel nomogram for predicting the prolonged length of stay in post-anesthesia care unit after elective operation.

BACKGROUND: Prolonged length of stay in post-anesthesia care unit (PLOS in PACU) is a combination of risk factors and complications that can compromise quality of care and operating room efficiency. Our study aimed to develop a nomogram to predict PLOS in PACU of patients undergoing elective surgery. METHODS: Data from 24017 patients were collected. Least absolute shrinkage and selection operator (LASSO) was used to screen variables. A logistic regression model was built on variables determined by a combined method of forward selection and backward elimination. Nomogram was designed with the model. The nomogram performance was evaluated with the area under the receiver operating characteristic curve (AUC) for discrimination, calibration plot for consistency between predictions and actuality, and decision curve analysis (DCA) for clinical application value. RESULTS: A nomogram was established based on the selected ten variables, including age, BMI < 21 kg/m2, American society of Anesthesiologists Physical Status (ASA), surgery type, chill, delirium, pain, naloxone, operation duration and blood transfusion. The C-index value was 0.773 [95% confidence interval (CI) = 0.765 - 0.781] in the development set and 0.757 (95% CI = 0.744-0.770) in the validation set. The AUC was > 0.75 for the prediction of PLOS in PACU. The calibration curves revealed high consistencies between the predicted and actual probability. The DCA showed that if the threshold probability is over 10% , using the models to predict PLOS in PACU and implement intervention adds more benefit. CONCLUSIONS: This study presented a nomogram to facilitate individualized prediction of PLOS in PACU for patients undergoing elective surgery.

Amplified Drug Delivery System with a Pair of Master Keys Triggering Precise Drug Release for Chemo-Photothermal Therapy.

A versatile drug delivery system (DDS) enabling highly effective and targeting oncotherapy has always been of great significance in medical research. In the development of a stimuli-responsive DDS, compared with a single-factor stimulation DDS, a multifactor activation DDS has higher therapeutic specificity between diseased and normal tissue, but there are challenges in drug-release efficiency and united targeting cancer therapy. Herein, a novel dual-microRNA (dual-miRNA)-mediated 1:N-amplified DDS is fabricated. The gold nanocage (AuNC) was synthesized and used as a carrier. A DNA bridge motif as a nanolock (DNA bridge nanolock) was designed and modified on the surface of AuNCs, which could seal the holes of AuNCs. Using the dual-miRNAs as a pair of master keys, through DNA strand migration and DNAzyme self-assembly, a cell endogenous substance Mg2+-dependent DNAzyme cyclic shear reaction could perform the function of the master keys to open multiple locks for the enhanced release of doxorubicin from the AuNCs. In addition, under near-infrared irradiation, via absorption of light and heat release, the AuNC is activated to perform the function of photothermal therapy. Thereby, the system achieves precise chemo-photothermal therapy. Using the in vitro and in vivo anti-tumor analysis, the DDS could be proved to present a novel design of enhanced and targeted drug-release system for highly effective cancer therapy.

KDM2A plays a dual role in regulating the expression of malignancy-related genes in esophageal squamous cell carcinoma.

KDM2A is a histone demethylase, which primarily catalyzes the demethylation of H3K36me2. Abnormal expression of KDM2A is observed in many types of cancers; however, the molecular events connected to KDM2A expression remain unclear. We report that KDM2A performs an oncogenic function in esophageal squamous cell carcinoma (ESCC) and is robustly expressed in ESCC cells. ShRNA-mediated knockdown of KDM2A resulted in a significant inhibition of the malignant phenotype of ESCC cell lines, whereas ectopic expression of KDM2A showed the opposite effect. We also analyzed the function of KDM2A using a CRISPR-CAS9 depletion system and subsequent rescue experiment, which also indicated a cancerous role of KDM2A. Interestingly, analysis of the gene expression network controlled by KDM2A using RNA-seq revealed an unexpected feature: KDM2A could induce expression of a set of well-documented oncogenic genes, including IL6 and LAT2, while simultaneously suppressing another set of oncogenes, including MAT2A and HMGCS1. Targeted inhibition of the upregulated oncogene in the KDM2A-depleted cells led to a synergistic suppressive effect on the malignant phenotype of ESCC cells. Our results revealed the dual role of KDM2A in ESCC cells, which may have therapeutic implications.

Fluorescent-Raman Binary Star Ratio Probe for MicroRNA Detection and Imaging in Living Cells.

The expression of microRNAs (miRNAs) is critical in gene regulation and has been counted into disease diagnosis marks. Precise imaging and quantification of miRNAs could afford the important information for clinical diagnosis. Here, two smart binary star ratio (BSR) probes were designed and constructed, and miRNA triggered the connection of the binary star probes and the reciprocal changes of dual signals in living cells. This multifunctional probe integrates fluorescence and surface enhanced Raman scattering (SERS) imaging, with enzyme-free numerator signal amplification for dual-mode imaging and dual-signal quantitative analysis of miRNA. First, compared with the single-mode ratio imaging method, using fluorescence-SERS complementary ratio imaging, this probe enables more accurate imaging contrast for direct visualization signal changes in living cells. Multiscale information about the dynamic behavior of miRNA and the probe is acquired. Next, via SERS reverse signal ratio response and a novel enzyme-free numerator signal amplification, the amplified signal and reduced black value were achieved in the quantification of miRNA. More importantly, BSR probes showed good stability in cells and were successfully used for accurate tracing and quantification of miR-203 from MCF-7 cells. Therefore, the reported BSR probe is a potential tool for the reliable monitoring of biomolecule dynamics in living cells.

Triggerable Mutually Amplified Signal Probe Based SERS-Microfluidics Platform for the Efficient Enrichment and Quantitative Detection of miRNA.

Sensitive detection of microRNAs (miRNAs) that serve as a disease marker could advance the diagnosis and treatment of diseases. Many methods used for quantitative detection of miRNAs, such as PCR-based approaches or the hybridization chain reaction, have presented challenges due to the complicated and time-consuming-procedures that are required. In this manuscript, a simple triggerable mutually amplified signal (TMAS) probe was designed and enriched within the center of a microfluidic chip and then used for one-step quantitative detection of microRNAs via surface enhanced Raman scattering (SERS) technology. First, many mutually amplified double strands are produced via an enzyme-free target-strand displacement recycling reaction initiated by the target miRNA, that result in the generation of an enhanced SERS signal. Second, microfluidic chips that utilize alternating current (AC) electrokinetic flow technology produce efficient mixing and rapid concentration to improve the DNA hybridization rate and further enhance the SERS signal intensity. This method enables the sensitive and rapid detection of miR-21 in human breast cancer cells within 30 min with a detection limit of 2.33 fM. Compared with traditional methods, this novel method overcomes the shortcomings resulting from complex operations, and has the advantages of high sensitivity, short assay time, and reduced sample usage.

Correction to: The effects and mechanisms of SLC34A2 in tumorigenesis and progression of human non-small cell lung cancer.

AbstractAfter the publication of this article [1] it came to our attention that there were some errors in two of the figures.

Prospective investigation of intravenous patient-controlled analgesia with hydromorphone or sufentanil: impact on mood, opioid adverse effects, and recovery.

BACKGROUND: Radical surgery for colorectal cancer, associated with moderate to severe postoperative pain, needs multimodal analgesia with opioid for analgesia. Despite considerable advancements, the psychological implications and other side effects with opioid remain substantially unresolved. This study aimed to investigate the impact on mood, side effects relative to opioid, and recovery of the patients with hydromorphone or sufentanil intravenous patient-controlled analgesia (IV-PCA) in a multimodal perioperative analgesia regimen undergoing radical surgery for colorectal cancer. METHODS: Eighty patients undergoing elective laparoscopic or open radical surgery for colorectal cancer under general anesthesia were randomized to receive postoperative IV-PCA with either sufentanil (group S) or hydromorphone (group H). All patients received additionally flurbiprofen axetil 50 mg 30 min before the end of surgery and wound infiltration with 10 ml of 0.75% ropivacaine at the end of surgery. The primary endpoint was mood changes at 48 and 96 h after surgery. The secondary endpoints were the incidence of opioid-related adverse effects, recovery results and patient satisfaction after surgery. RESULTS: Seventy-two patients completed the study finally. There were no significant differences between the two groups with respect to preoperative parameters, surgical and anesthetic characteristics (P > 0.05). No obvious significant differences were observed in VAS score (at rest and during mobilization) and rescue analgesics use (P > 0.05). Compared with group S, the anger scores in the group H at 48 h and 96 h after surgery were significantly lower (P = 0.012 and 0.005; respectively), but the incidences of pruritus and nausea were higher (P = 0.028 and 0.008; respectively). There were no significant differences in the incidences of vomiting, respiratory depression, dizziness, Ramsay score, and hemodynamic changes between the two groups (P > 0.05). Moreover, there were no significant differences in the time to gastrointestinal recovery, time to drainage tube removal, time to walk, hospital stay after surgery and patient satisfaction between the two groups (P > 0.05). CONCLUSIONS: Under the similar analgesia effect with different opoiods postoperatively, hydromorphone IV-PCA resulted in an improved mood, however, a higher occurrence of pruritus and nausea while compared to sufentanil IV-PCA in a multimodal perioperative analgesia regimen. Both regimens of opioid with IV-PCA may serve as promising candidates for good postoperative pain management, and provide with similar postoperative recovery for the patients undergoing radical surgery for colorectal cancer. TRIAL REGISTRATION: This study was registered with the Chinese Clinical Trial Registry on September 20, 2015 (URL: http://www.chictr.org.cn . Registry number: ChiCTR-IPR-15007112).

Fluorescence and SERS Imaging for the Simultaneous Absolute Quantification of Multiple miRNAs in Living Cells.

The simultaneous imaging and quantification of multiple intracellular microRNAs (miRNAs) are particularly desirable for the early diagnosis of cancers. However, simultaneous direct imaging with absolute quantification of multiple intracellular RNAs remains a great challenge, particularly for miRNAs, which have significantly different expression levels in living cells. We designed dual-signal switchable (DSS) nanoprobes using the fluorescence-Raman signal switch. The intracellular uptake and dynamic behaviors of the probe are monitored by its fluorescence signal. Meanwhile, real-time quantitative detection of multiple miRNAs is made possible by measurements of the surface-enhanced Raman spectroscopy (SERS) ratios. Moreover, the signal 1:n ratio amplification mode only responds to low-abundance miRNA (asymmetric signal amplification mode) for simultaneous visualization and quantitative detection of significantly different levels of miRNAs in living cells. miR-21 and miR-203 were successfully detected in living MCF-7 cells, in agreement with in vitro results from the same batch of cell lysates. The reported dual-spectrum imaging method promises to offer a new strategy for the intracellular imaging and detection of various types of biomolecules.

Asymmetric signal amplification for simultaneous SERS detection of multiple cancer markers with significantly different levels.

Simultaneous detection of cancer biomarkers holds great promise for the early diagnosis of different cancers. However, in the presence of high-concentration biomarkers, the signals of lower-expression biomarkers are overlapped. Existing techniques are not suitable for simultaneously detecting multiple biomarkers at concentrations with significantly different orders of magnitude. Here, we propose an asymmetric signal amplification method for simultaneously detecting multiple biomarkers with significantly different levels. Using the bifunctional probe, a linear amplification mode responds to high-concentration markers, and quadratic amplification mode responds to low-concentration markers. With the combined biobarcode probe and hybridization chain reaction (HCR) amplification method, the detection limits of microRNA (miRNA) and ATP via surface-enhanced Raman scattering (SERS) detection are 0.15 fM and 20 nM, respectively, with a breakthrough of detection concentration difference over 11 orders of magnitude. Furthermore, successful determination of miRNA and ATP in cancer cells supports the practicability of the assay. This methodology promises to open an exciting new avenue for the detection of various types of biomolecules.

The effects and mechanisms of SLC34A2 in tumorigenesis and progression of human non-small cell lung cancer.

BACKGROUND: SLC34A2 with highest expressions in lung, small intestine and kidney encoded a type 2b sodium-dependent phosphate transporter (NaPi-IIb). In lung, SLC34A2 only expressed in the apical membrane of type II alveolar epithelium cells (ATII cells) and played a pivotal role during the fetal lung development and embryonic development. ATII cells acting as multifunctional stem cells might transform into NSCLC after undergoing exogenous or endogenous factors. Increasing evidences showed that the genes performing critical roles during embryogenesis were also expressed during the development of cancer. In addition, recent research found the expression of SLC34A2 had a significant difference between the surgical samples of NSCLC and normal tissues, and SLC34A2 was down-regulated in lung adenocarcinoma cell line A549 and up-regulation expression of SLC34A2 could significantly inhibit cell viability and invasion of A549 in vitro. These results suggested SLC34A2 might play an important role in the development of NSCLC. However, the role of SLC34A2 in tumorigenesis and progression of NSCLC remains unknown. RESULTS: Our study found that SLC34A2 was also significantly down-regulated in 14/15 of examined NSCLC tissues. Moreover, we found that expressions of SLC34A2 were reduced in six NSCLC cell lines for the first time. Our result also revealed a dramatic inhibitory effects of SLC34A2 on cell growth, migration and invasion of several NSCLC cell lines. SLC34A2 also strongly inhibited tumor growth and metastasis ability in A549 subcutaneous tumor model and lung metastasis model, respectively. Further studies found that the suppressive effects of SLC34A2 on tumorigenesis and progression might be associated with the down-regulation of related protein in PI3K/Akt and Ras/Raf/MEK signal pathway. CONCLUSIONS: For the first time, our data indicated that SLC34A2 could exert significantly suppressive effects on tumorigenesis and progression of NSCLC. SLC34A2 might provide new insights for further understanding the early pathogenesis of human NSCLC.

Aptamer-based SERS assay of ATP and lysozyme by using primer self-generation.

A simple bifunctional surface-enhanced Raman scattering (SERS) assay based on primer self-generation strand-displacement polymerization (PS-SDP) is developed to detect small molecules or proteins in parallel. Triphosphate (ATP) and lysozyme are used as the models of small molecules and proteins. Compared to traditional bifunctional methods, the method possesses some remarkable features as follows: 1) by virtue of the simple PS-SDP reaction, a bifunctional aptamer assembly binding of trigger 1 and trigger 2 was used as a functional structure for the simultaneous sensing of ATP or lysozyme. 2) The concept of isothermal amplification bifunctional detection has been first introduced into SERS biosensing applications as a signal-amplification tool. 3) The problem of high background induced by excess bio-barcodes is circumvented by using magnetic beads (MBs) as the carrier of signal-output products and massive of hairpin DNA binding with SERS active bio-barcodes relied on Au nanoparticles (Au NPs), SERS signal is significantly enhanced. Overall, with multiple amplification steps and one magnetic-separation procedure, this flexible biosensing system exhibited not only high sensitivity and specificity, with the detection limits of ATP and lysozyme of 0.05 nM and 10 fM, respectively.

Efficacy and safety of anlotinib plus XELOX regimen as first-line therapy for mCRC: a single-arm, multicenter, phase II study (ALTER-C-001).

Background: Anlotinib showed encouraging anti-tumor activity in metastatic colorectal cancer (mCRC). This study was designed to assess the efficacy and safety of anlotinib plus XELOX as first-line therapy in mCRC patients. Materials and Methods: Eligible patients aged ≥18 with mCRC were enrolled in this multicenter, single-arm, phase II, exploratory study. Patients received at least 6 cycles of anlotinib, oxaliplatin, and capecitabine as initial therapy. Subsequently, patients received anlotinib monotherapy as maintenance therapy until tumor progression or intolerable toxicity. The primary endpoint was progression-free survival (PFS). Results: Thirty-one patients were included between December 2019 and March 2022. The median follow-up was 17.5 (95% CI, 3.0-17.5) months. The median PFS was 8.3 (95% CI, 6.3-10.0) months, with 6- and 12-month PFS rates of 82.3% (95% CI, 59.2%-93.0%) and 18.9% (95% CI, 4.8%-40.1%), respectively. Fifteen (48.4%) achieved partial response for an ORR of 48.4% (95% CI, 30.2%-66.9%). The disease control rate was 71.0% (95% CI, 52.0%-85.8%) due to 7 (22.6%) stable diseases. The median duration of response was 6.0 (95% CI, 3.6-8.0) months and 1 patient had the longest ongoing response of 17.3 months. Of 24 patients with evaluable imaging, 23 (74.2%) obtained tumor shrinkage. The median PFS (11.0 vs. 6.9 months) and ORR (66.7% vs. 60.0%) for patients with RAS/BRAF wild-type were numerically better than those with mutation. Three patients are still ongoing treatment. The grade 3 or more treatment-emergent adverse events (TEAEs) were mainly hypertension (12.9%) and decreased neutrophil count (12.9%). Four (12.9%) had serious TEAEs, primarily including abdominal pain and incomplete intestinal obstruction. Conclusion: Anlotinib plus XELOX as first-line therapy in patients with mCRC showed anti-tumor activity and safety profile, which is worth further investigation. Clinical Trial Registration: chictr.org.cn, identifier ChiCTR1900028417.

A new signal-on photoelectrochemical biosensor based on a graphene/quantum-dot nanocomposite amplified by the dual-quenched effect of bipyridinium relay and AuNPs.

A new photoelectrochemical (PEC) biosensor was developed by using carboxyl-functionalized graphene and CdSe nanoparticles. This sensitive interface was then successfully applied to detection of thrombin based on the dual-quenched effect of PEC nanoparticle, which relied on the electron transfer of a bipyridinium relay and energy transfer of AuNPs. After recognition with an aptamer, the PEC nanoparticle was removed and a signal-on PEC biosensor was obtained. Moreover, the bio-barcode technique used in the preparation of PEC nanoparticle could avoid cross-reaction and enhances the sensitivity. Taking advantages of the various methods mentioned above, the sensitivity could be easily enhanced. In addition, in this work we also investigated graphene that was modified with different functional groups and AuNPs of different particle sizes. Under optimal conditions, a detection limit of 5.9×10(-15) M was achieved. With its simplicity, selectivity, and sensitivity, this strategy shows great promise for the fabrication of highly efficient PEC biosensors.

SERS-Microfluidic Approach for the Quantitative Detection of miRNA Using DNAzyme-Mediated Reciprocal Signal Amplification.

MicroRNAs (miRNAs) play important roles in biological processes. Designing a sensitive, selective, and rapid method of miRNA detection is crucial for biological research. Here, with a reciprocal signal amplification (RSA) probe, this work established a novel surface-enhanced Raman scattering (SERS)-microfluidic approach for the quantitative analysis of miRNA. First, via a DNAzyme self-assemble cycle reaction, two types of SERS signals produce amplified reciprocal changes. The sum of the absolute signal value is first adopted for the quantitative analysis of miRNA, which results in an enhanced response and a reduced blank value. Furthermore, the assay is integrated in an electric drive microfluidic mixing reactor that enables physical mixing and enriching of the reactants for more rapid and enhanced detection sensitivity. The protocol owns the merits of the SERS technology, amplified reciprocal signals, and a microfluidic chip, with a detection limit of 2.92 fM for miR-141 in 40 min. In addition, successful determination of miRNA in a variety of cells proved the practicability of the assay. Compared with the reported strategies for miRNA analysis, this work avoids a complex and time-consuming procedure and enhances the sensitivity and specificity. The method opens a promising way for biomolecular chip detection and research.

A One-Two-Three Multifunctional System for Enhanced Imaging and Detection of Intracellular MicroRNA and Chemogene Therapy.

Simultaneous imaging, diagnosis, and therapy can offer an effective strategy for cancer treatment. However, the complex probe design, poor drug release efficiency, and multidrug resistance remain tremendous challenges to cancer treatment. Here, a novel one-two-three system is built for enhanced imaging and detection of miRNA-21 (miR-21) overexpressed in cancer cell and chemogene therapy. The system consists of dual-mode DNA robot nanoprobes assembled by two types of hairpin DNAs and three-way branch DNAs modified on gold nanoparticles, with intercalating anticancer drugs (doxorubicin), into DNA duplex GC base pairs. In the system, via intracellular ATP-accelerated cyclic reaction triggered by miR-21, fluorescence and SERS signals were alternated with DNA structure switch, and the precise SERS detection of miRNA and fluorescence imaging oriented "on-demand" release of two types of anticancer drugs (anti-miR-21 and Dox) are achieved. Thus, "one-two-three" means one kind of miR-21-triggered endogenous substance accelerated cyclic reaction, two modes of signal switch, and three functions including enhanced imaging, detection, and comprehensive treatment. The one-two-three system has some notable merits. First, ATP as an endogenous substance promotes DNA structure switching and accelerates the cyclic reaction. Second, the treatment with a dual-mode signal switch is more reliable and accurate and can provide more abundant information than a single-mode treatment platform. Thus, the imaging and detection of intracellular miRNA and effective comprehensive therapy are realized. In vivo results indicate that the system can provide new insights and strategies for diagnosis and therapy.

Multiple-mRNA-controlled and heat-driven drug release from gold nanocages in targeted chemo-photothermal therapy for tumors.

Multifunctional drug delivery systems enabling effective drug delivery and comprehensive treatment are critical to successful cancer treatment. Overcoming nonspecific release and off-target effects remains challenging in precise drug delivery. Here, we design triple-interlocked drug delivery systems to perform specific cancer cell recognition, controlled drug release and effective comprehensive therapy. Gold nanocages (AuNCs) comprise a novel class of nanostructures possessing hollow interiors and porous walls. AuNCs are employed as a drug carrier and photothermal transducer due to their unique structure and photothermal properties. A smart triple-interlocked I-type DNA nanostructure is modified on the surface of the AuNCs, and molecules of the anticancer drug doxorubicin (DOX) are loaded as molecular cargo and blocked. The triple-interlocked nanostructure can be unlocked by binding with three types of tumor-related mRNAs, which act as "keys" to the triple locks, sequentially, which leads to precise drug release. Additionally, fluorescence-imaging-oriented chemical-photothermal synergistic treatment is achieved under illumination with infrared light. This drug delivery system, which combines the advantages of AuNCs and interlocked I-type DNA, successfully demonstrates effective and precise imaging, drug release and photothermal therapy. This multifunctional triple-interlocked drug delivery system could be used as a potential carrier for effective cancer-targeting comprehensive chemotherapy and photothermal therapy treatments.

[Study on invasion and metastasis-associated genes of lung cancer related with NM23-H1 gene].

OBJECTIVE: To study on invasion and metastasis-associated genes of lung cancer related with NM23-H1 gene. METHODS: Human gene expression chip based on the subtracted cDNA libraries was constructed. After microarray hybridization, clones sequencing, sequence homology search, the information of differently expressed genes in human large cell lung cancer cell line of L9981 and L9981-nm23-H1 were obtained and then further confirmed by real-time quantitative PCR. RESULTS: Gene expression profiling chips of differently expressed genes in human large cell lung cancer cell line L9981 and L9981-nm23-H1 were successfully constructed. After microarray hybridization, sequence homology search, 19 differentially expressed genes were observed. After real-time quantitative PCR evaluation, we found that the mRNA of 8 genes including PSMA7, SBDS, ODC1, YARS, CSDA, PTP4A1, SHPRH and TOMM7 was up-regulated in the cell line of L9981 after transfected with NM23-H1 gene, whereas the mRNA of PKM2 and GMNN was down-regulated. CONCLUSION: NM23-H1 gene may be the upstream regulator of metastasis-associated genes, which can regulate the downstream genes to achieve a series of lung cancer metastatic potential.

An accurate and ultrasensitive SERS sensor with Au-Se interface for bioimaging and in situ quantitation.

We propose a stable and highly sensitive Au-Se SERS nanoprobe for bioimaging and in situ quantitation, which aims to break through the limitations of traditional Au-S SERS nanoprobes, such as interference from biothiols and unsatisfactory SERS efficiency.

Analysis of Saccharomyces cerevisiae genome for the distributions of stress-response elements potentially affecting gene expression by transcriptional interference.

Cellular stress responses are characterized by coordinated transcriptional induction of genes encoding a group of conserved proteins known as molecular chaperones, most of which are also known as heat shock proteins (HSPs). In S. cerevisiae, transcriptional responses to stress are mediated via two trans-regulatory activators: heat shock transcription factors (HSFs) that bind to heat shock elements (HSEs), and the Msn2 and Msn4 transcription factors that bind to stress response elements (STREs). Recent studies in S. cerevisiae demonstrated that a significant portion of the non-coding region in the genome is transcribed and this intergenic transcription could regulate the transcription of adjacent genes by transcription interference. The goal of this study was to analyze the genomic distribution of HSF and Msn2/4 binding sites and to study the potential for transcription interference regulated by stress response systems. Our genome-wide analysis revealed that 297 genes have STREs in their promoter region, whereas 310 genes contained HSEs. Twenty-five genes had both HSEs and STREs in their promoters. The first set of genes is potentially regulated by the Msn2/Msn4/STRE interaction. For the second set of genes, regulation by heat shock could be mediated through HSF/HSE regulatory mechanisms. The overlap between these groups suggests a co-regulation by the two pathways. Our study yielded 239 candidate genes, whose regulation could potentially be affected by heat-shock via transcription interference directed both from upstream and downstream areas relative to the native promoters. In addition we have categorized 924 genes containing HSE and/or STRE elements within the Open Reading Frames (ORFs), which may also affect normal transcription. Our study revealed a widespread possibility for the regulation of genes via transcriptional interference initiated by stress response. We provided a categorization of genes potentially affected at the transcriptional level by known stress-response systems.